Isolation of Active Regulatory Elements from Eukaryotic Chromatin Using FAIRE (Formaldehyde-Assisted Isolation of Regulatory Elements)

Department of Biology and Carolina Center for the Genome Sciences, University of North Carolina at Chapel Hill, CB #3280, 408 Fordham Hall, Chapel Hill, NC 27599-3280, USA
Methods (Impact Factor: 3.22). 07/2009; 48(3):233-239. DOI: 10.1016/j.ymeth.2009.03.003

ABSTRACT The binding of sequence-specific regulatory factors and the recruitment of chromatin remodeling activities cause nucleosomes to be evicted from chromatin in eukaryotic cells. Traditionally, these active sites have been identified experimentally through their sensitivity to nucleases. Here we describe the details of a simple procedure for the genome-wide isolation of nucleosome-depleted DNA from human chromatin, termed FAIRE (Formaldehyde Assisted Isolation of Regulatory Elements). We also provide protocols for different methods of detecting FAIRE-enriched DNA, including use of PCR, DNA microarrays, and next-generation sequencing. FAIRE works on all eukaryotic chromatin tested to date. To perform FAIRE, chromatin is crosslinked with formaldehyde, sheared by sonication, and phenol–chloroform extracted. Most genomic DNA is crosslinked to nucleosomes and is sequestered to the interphase, whereas DNA recovered in the aqueous phase corresponds to nucleosome-depleted regions of the genome. The isolated regions are largely coincident with the location of DNaseI hypersensitive sites, transcriptional start sites, enhancers, insulators, and active promoters. Given its speed and simplicity, FAIRE has utility in establishing chromatin profiles of diverse cell types in health and disease, isolating DNA regulatory elements en masse for further characterization, and as a screening assay for the effects of small molecules on chromatin organization.

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    • "Like most other transcription factors, VDR competes with the intrinsic repressive nature of chromatin for access to its genomic binding sites [25] [26]. Open chromatin regions can be detected by the genome-wide method formaldehyde-assisted isolation of regulatory elements sequencing (FAIRE-seq) [27] [28] [29]. "
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    • "In contrast to a recent study showing that SIRT7 deacetylates H3K18 at promoters of ELK4 target genes (Barber et al., 2012), RNAi-mediated depletion of SIRT7 did not affect H3K18 acetylation at rDNA (Figure S2B), suggesting that SIRT7 deacetylates different substrates in a gene-specific context. Moreover, Formaldehyde Assisted Isolation of Regulatory Elements (FAIRE) assays which measure nucleosome density according to extractability of crosslinked chromatin (Giresi and Lieb, 2009) revealed no changes in rDNA chromatin compaction, supporting the notion that SIRT7 activates transcription by targeting the Pol I transcription apparatus rather than by changing the chromatin structure (Figure 2G). "
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    • "FAIRE analysis was performed according to the protocol published by Giresi et al. [27]. In short, 18 × 10 6 THP-1 cells were cross-linked identically as for ChIP. "
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